NZ765451B2 - Antisense nucleic acid - Google Patents

Antisense nucleic acid

Info

Publication number
NZ765451B2
NZ765451B2 NZ765451A NZ76545115A NZ765451B2 NZ 765451 B2 NZ765451 B2 NZ 765451B2 NZ 765451 A NZ765451 A NZ 765451A NZ 76545115 A NZ76545115 A NZ 76545115A NZ 765451 B2 NZ765451 B2 NZ 765451B2
Authority
NZ
New Zealand
Prior art keywords
nucleotide sequence
antisense oligomer
seq
pharmaceutically acceptable
oligomer
Prior art date
Application number
NZ765451A
Other versions
NZ765451A (en
Inventor
Tetsuya Nagata
Shin Ichi Takeda
Yuuichirou TONE
Naoki Watanabe
Original Assignee
National Center Of Neurology And Psychiatry
Nippon Shinyaku Co Ltd
Filing date
Publication date
Application filed by National Center Of Neurology And Psychiatry, Nippon Shinyaku Co Ltd filed Critical National Center Of Neurology And Psychiatry
Publication of NZ765451A publication Critical patent/NZ765451A/en
Publication of NZ765451B2 publication Critical patent/NZ765451B2/en

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Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/7088—Compounds having three or more nucleosides or nucleotides
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/7088—Compounds having three or more nucleosides or nucleotides
    • A61K31/712—Nucleic acids or oligonucleotides having modified sugars, i.e. other than ribose or 2'-deoxyribose
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/7088—Compounds having three or more nucleosides or nucleotides
    • A61K31/7125—Nucleic acids or oligonucleotides having modified internucleoside linkage, i.e. other than 3'-5' phosphodiesters
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K48/00—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
    • C12N15/113—Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/10—Type of nucleic acid
    • C12N2310/11—Antisense
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/30—Chemical structure
    • C12N2310/32—Chemical structure of the sugar
    • C12N2310/323—Chemical structure of the sugar modified ring structure
    • C12N2310/3233—Morpholino-type ring
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2320/00—Applications; Uses
    • C12N2320/10—Applications; Uses in screening processes
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2320/00—Applications; Uses
    • C12N2320/30—Special therapeutic applications
    • C12N2320/33—Alteration of splicing
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2330/00—Production
    • C12N2330/30—Production chemically synthesised
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q2600/00—Oligonucleotides characterized by their use
    • C12Q2600/136—Screening for pharmacological compounds
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q2600/00—Oligonucleotides characterized by their use
    • C12Q2600/178—Oligonucleotides characterized by their use miRNA, siRNA or ncRNA

Abstract

Provided is a drug that allows highly-efficient skipping of exon. The present invention provides an antisense oligomer wherein two or more unit oligomers targeting sequences that are neither consecutive nor overlap with each other in the same exon are connected.

Claims (24)

Claims:
1. An antisense oligomer having a length of 15 to 30 bases, consisting of a first unit oligomer and a second unit oligomer which are connected, wherein (a) the first unit oligomer consisting of a nucleotide sequence complementary to a first nucleotide sequence of 7 to 15 consecutive bases in exon 44 in human dystrophin gene; (b) the second unit oligomer consisting of a nucleotide sequence complementary to a second nucleotide sequence of 7 to 15 consecutive bases in exon 44, wherein the first nucleotide sequence and the second nucleotide sequence are neither consecutive nor overlap with each other, and wherein the antisense oligomer induces skipping of exon 44, wherein the first nucleotide sequence is a nucleotide sequence of 7 to 15 consecutive bases selected from the nucleotide sequence represented by SEQ ID NO: 1 or SEQ ID NO:2 and the second nucleotide sequence is a nucleotide sequence of 7 to 15 consecutive bases selected from the nucleotide sequence represented by SEQ ID NO: 2, or a pharmaceutically acceptable salt or hydrate thereof.
2. The antisense oligomer according to claim 1, wherein the first nucleotide sequence is a nucleotide sequence of 7 to 15 consecutive bases selected from the nucleotide sequence represented by SEQ ID NO: 1, or a pharmaceutically acceptable salt or hydrate thereof.
3. The antisense oligomer according to claim 1 or 2, wherein the first nucleotide sequence or second nucleotide sequence is a nucleotide sequence of 7 to 15 consecutive bases selected from the nucleotide sequence represented by SEQ ID NO: 2, or a pharmaceutically acceptable salt or hydrate thereof.
4. An antisense oligomer having a length of 15 to 30 bases, wherein two unit oligomers selected from the group consisting of the following (c) to (e) are connected: (c) a unit oligomer consisting of a nucleotide sequence complementary to a nucleotide sequence of 7 to 15 consecutive bases selected from the nucleotide sequence represented by SEQ ID NO: 3; (d) a unit oligomer consisting of a nucleotide sequence complementary to a nucleotide sequence of 7 to 15 consecutive bases selected from the nucleotide sequence represented by SEQ ID NO: 4; and (e) a unit oligomer consisting of a nucleotide sequence complementary to a nucleotide sequence of 7 to 15 consecutive bases selected from the nucleotide sequence represented by SEQ ID NO: 5, wherein the two nucleotide sequences are neither consecutive nor overlap with each other, and wherein the antisense oligomer induces skipping of exon 44, or a pharmaceutically acceptable salt or hydrate thereof.
5. The antisense oligomer according to claim 1, consisting of a nucleotide sequence set forth in SEQ ID NO: 6.
6. The antisense oligomer according to claim 1, consisting of a nucleotide sequence set forth in SEQ ID NO: 7.
7. The antisense oligomer according to claim 1, consisting of a nucleotide sequence set forth in SEQ ID NO: 8.
8. The antisense oligomer according to claim 1, consisting of a nucleotide sequence set forth in SEQ ID NO: 9.
9. The antisense oligomer according to claim 1, consisting of a nucleotide sequence set forth in SEQ ID NO: 55.
10. The antisense oligomer according to claim 1, consisting of a nucleotide sequence set forth in SEQ ID NO: 106.
11. The antisense oligomer according to any one of claims 1 to 10, which is an oligonucleotide, or a pharmaceutically acceptable salt or hydrate thereof.
12. The antisense oligomer according to claim 11, wherein the sugar moiety and/or the phosphate-binding region of at least one nucleotide constituting the oligonucleotide is modified, or a pharmaceutically acceptable salt or hydrate thereof.
13. The antisense oligomer according to claim 11 or 12, wherein the sugar moiety of at least one nucleotide constituting the oligonucleotide is a ribose in which the 2'-OH group is replaced by any one selected from the group consisting of OR, R, R'OR, SH, SR, NH , NHR, NR , N , CN, F, Cl, Br and I (wherein R is an alkyl or an aryl and R' is an 2 2 3 alkylene), or a pharmaceutically acceptable salt or hydrate thereof.
14. The antisense oligomer according to any one of claims 11 to 13, wherein the phosphate-binding region of at least one nucleotide constituting the oligonucleotide is any one selected from the group consisting of a phosphorothioate bond, a phosphorodithioate bond, an alkylphosphonate bond, a phosphoramidate bond and a boranophosphate bond, or a pharmaceutically acceptable salt or hydrate thereof.
15. The antisense oligomer according to any one of claims 1 to 10, which is a morpholino oligomer, or a pharmaceutically acceptable salt or hydrate thereof.
16. The antisense oligomer according to claim 15, which is a phosphorodiamidate morpholino oligomer, or a pharmaceutically acceptable salt or hydrate thereof.
17. The antisense oligomer according to claim 15 or 16, wherein the 5' end is any one of chemical formulae (1) to (3) below, or a pharmaceutically acceptable salt or hydrate thereof.
18. A pharmaceutical composition for the treatment of muscular dystrophy, comprising as an active ingredient the antisense oligomer according to any one of claims 1 to 17, or a pharmaceutically acceptable salt or hydrate thereof.
19. The pharmaceutical composition according to claim 18, comprising a pharmaceutically acceptable carrier.
20. Use of the antisense oligomer or a pharmaceutically acceptable salt or hydrate thereof according to any one of claims 1 to 17 in manufacturing a pharmaceutical composition for the treatment of muscular dystrophy in a patient.
21. The use according to claim 20, wherein the patient with muscular dystrophy has a mutation(s) which is to be targeted for exon 44 skipping in dystrophin gene.
22. A method for manufacturing of the antisense oligomer according to any one of claims 1 to 17, which comprises connecting (a) a first unit oligomer consisting of a nucleotide sequence complementary to a first nucleotide sequence of 7 to 15 consecutive bases in exon 44 in human dystrophin gene; (b) a second unit oligomer consisting of a nucleotide sequence complementary to a second nucleotide sequence of 7 to 15 consecutive bases in exon 44 to produce an antisense oligomer having a length of 15 to 30 bases, wherein the first nucleotide sequence and the second nucleotide sequence are neither consecutive nor overlap with each other wherein the first nucleotide sequence is a nucleotide sequence of 7 to 15 consecutive bases selected from the nucleotide sequence represented by SEQ ID NO: 1 or SEQ ID NO:2 and the second nucleotide sequence is a nucleotide sequence of 7 to 15 consecutive bases selected from the nucleotide sequence represented by SEQ ID NO: 2.
23. The method according to claim 22, which further comprises: measuring the efficiency of skipping by the obtained antisense oligomer; and selecting an antisense oligomer having the efficiency of skipping that exceeds a reference value.
24. A method for screening of an antisense oligomer, which comprises: (a) selecting (i) a first unit oligomer consisting of a nucleotide sequence complementary to a first nucleotide sequence of 7 to 15 consecutive bases in exon 44 in human dystrophin gene; (ii) a second unit oligomer consisting of a nucleotide sequence complementary to a second nucleotide sequence of 7 to 15 consecutive bases in exon 44, wherein the first nucleotide sequence and the second nucleotide sequence are neither consecutive nor overlap with each other; (b) connecting the first and second unit oligomers to produce an antisense oligomer having a length of 15 to 30 bases consisting of the first and second unit oligomers; (c) measuring the efficiency of skipping by the antisense oligomer obtained in the step (b); and (d) selecting an antisense oligomer having the efficiency of skipping that exceeds a reference value wherein the first nucleotide sequence is a nucleotide sequence of 7 to 15 consecutive bases selected from the nucleotide sequence represented by SEQ ID NO: 1 or SEQ ID NO:2 and the second nucleotide sequence is a nucleotide sequence of 7 to 15 consecutive bases selected from the nucleotide sequence represented by SEQ ID NO: 2. G1076WO_ST25.txt
NZ765451A 2015-06-16 Antisense nucleic acid NZ765451B2 (en)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
JP2014124157 2014-06-17
NZ728103A NZ728103B2 (en) 2014-06-17 2015-06-16 Antisense nucleic acids

Publications (2)

Publication Number Publication Date
NZ765451A NZ765451A (en) 2024-04-26
NZ765451B2 true NZ765451B2 (en) 2024-07-30

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